bd logoRhapsody Sequence Analysis Pipeline N22_WTA
Pipeline Version: 2.0
Tue Jan 02 2024 16:11:41 GMT+0000 (Greenwich Mean Time)

Summary

Library
Total Reads in FASTQ
Pct Reads Passing Quality Filter
Pct CellLabel UMI Aligned Uniquely
N22 WTA 113,058,879 61.55% 41.66%
Putative Cells

564

Bioproduct Type
Aligned Reads By Type
Mean Reads per Cell
Mean Molecules per Cell
mRNA 8,076,193 1,502.27 72

Graphs

Sequence Quality

Library
Total Reads in FASTQ
Pct Read Pair Overlap
Pct Reads Too Short
Pct Reads Low Base Quality
Pct Reads High SNF
Pct Reads Filtered Out
Total Reads After Quality Filtering
N22 WTA 113,058,879 0% 34.51% 0.08% 3.86% 38.45% 69,591,971

Library Quality

Library
Total Filtered Reads
Pct Q30 Bases in Filtered R2
Pct CellLabel UMI
Pct CellLabel UMI Aligned Uniquely
Pct Reads Useful
N22 WTA 69,591,971 80.91% 65.6% 41.66% 11.61%

Alignment Categories

Library
CellLabel UMI Reads
Annotated Transcriptome Pct
Introns Pct
Intergenic Regions Pct
Antisense Pct
Not Unique Pct
Ambiguous Pct
No Feature Pct
AbSeq Pct
Sample Tag Pct
VDJ TCR Pct
VDJ BCR Pct
Unaligned Pct
N22 WTA 45,650,407 4.9% 12.79% 15.75% 13.55% 15.48% 1% 0.03% 0% 0% 0% 0% 36.5%

Reads and Molecules

Bioproduct Type
Aligned Reads By Type
Total Raw Molecules
Total RSEC Molecules
Mean Raw Sequencing Depth
Mean RSEC Sequencing Depth
Sequencing Saturation
mRNA 8,076,193 3,299,718 3,164,204 2.45 2.55 70.13

Cells

Putative Cells


Basic Algorithm Cells
Bioproduct Type
Pct Reads from Putative Cells
Mean Reads per Cell
Median Reads per Cell
Mean Molecules per Cell
Median Molecules per Cell
Mean Bioproducts per Cell
Median Bioproducts per Cell
Total Bioproducts Detected
mRNA 10.49% 1,502.27 1,189 72 51 58.83 46 10,876

Pipeline Inputs

Pipeline version: 2.0

Pipeline Start Time: Tue Jan 02 2024 16:11:41 GMT+0000 (Greenwich Mean Time)

AbSeq Reference:
  • No value
Reference Files Archive:
  • RhapRef_Human_WTA_2023-02.tar.gz
Supplemental Reference:
  • No value
Targeted Reference:
  • No value
Reads:
  • N22_WTA_S1_R1_001.fastq.gz
  • N22_WTA_S1_R2_001.fastq.gz
Multiplexing Settings

Sample Tags Version: No value
Tag Names:

VDJ Settings

VDJ Species Version: No value

Putative Cell Calling Settings

Enable Refined Putative Cell Calling: False
Exact Cell Count: No value
Expected Cell Count: No value
Putative Cell Call: mRNA

Exclude Intronic reads: False

Generate Bam output: True

Metric Alerts

(Under development)
  For the [N22_WTA] library, the [Pct_Reads_Too_Short] metric value of 34.51% is greater than 5% (Sequencing Quality section, WTA assay threshold).
   
  • May be due to short products in the library prep.
  • Look for evidence of this in QC traces and contact customer support for suggestions on additional cleanups.
  For the [N22_WTA] library, the [Pct_CellLabel_UMI] metric value of 65.6% is less than 85% (Library Quality section, WTA V2 Bead assay threshold).
   
  • May be due to poor sequencing or library quality, or incorrect reference panel.
  • Check Q30 scores and QC traces, and confirm that the appropriate transcriptome reference and complete AbSeq reference was used.
  • This can also occur if V1 and V2 beads are combined in a single library.
  For the [N22_WTA] library, the [Unaligned_Pct] metric value of 36.5% is greater than 10% (Alignment Categories section, WTA assay threshold).
   
  • May occur in experiments with low cell input. This can be addressed with additional cleanups for low cell input libraries (included in the protocol).
  • Can also result from low cell viability or poor sequencing or library quality.
  • This is expected if samples such as nuclei containing a high proportion of unprocessed transcripts are being run.
  For the [mRNA] bioproduct type, the [Pct_Reads_from_Putative_Cells] metric value of 10.49% is less than 75% (Cells section, WTA assay threshold).
   
  • May be an indication of low cell viability going onto the cartridge.
  • Confirm on scanner that an appropriate proportion of live cells were captured.
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